To understand how the brain computes, we need to understand how individual neurons in a circuit integrate their numerous inputs into output signals, as well as how they work together to encode a sensory input or execute a motor command in a behaving animal. Circuits and neurons are three-dimensional (3D) and can extend over hundreds or thousands of microns. Therefore, understanding their operations requires monitoring their activity at both synaptic and cellular resolution in 3D at image rates that capture all activity events. Behaving animals present a host of challenges to this goal. Existing 3D imaging technologies suffer from insufficient volume imaging speed, brain-motion-induced image artifacts, as well as complex hardware and software implementation. These limitations have prevented their adoption by biology laboratories and remain a technical barrier for neuroscience research. Successful completion of our proposal will overcome these limitations and profoundly impact neuroscience research. We recently developed a Bessel focus scanning technology (BEST) that is easily integrated into existing two-photon microscopes, resistant to motion artifacts, and have already achieved 30-Hz, synapse-resolving volumetric imaging of sparsely labelled neuronal populations in a wide variety of model organisms. In this proposal, combining the expertise of microscopists, biologists, and data scientists, we propose to further optimize BEST to enable high-speed, high-throughput, and high-resolution volumetric activity recording of both sparsely and densely labelled circuits throughout the living brain. We aim to record whole-brain activity in the fly at >10 Hz and through-cortex volume imaging in the mouse at ~2Hz. By combining BEST with microendoscopy, we aim to achieve synaptic-resolution volumetric microendoscopic imaging at 30 Hz and use it to study structural and functional plasticity in deeply buried nuclei of the mouse brain. By correcting brain-induced optical aberrations, adaptive optics will enable BEST to maintain synapse resolution throughout the entire mouse cortex. Easily adoptable, BEST has already been integrated into multiple two-photon fluorescence microscopes in laboratories worldwide. With a continuously expanding user base, the proposed optimization project will immediately benefit a wide range of laboratories, allowing them to study volumetric neural activity at unprecedented high spatiotemporal resolution throughout the living brain.